Description: (Taken directly from the application) Hypocitraturia is an important cause of kidney stones. Citrate is freely filtered in the glomerulus, and urinary citrate excretion is regulated principally by the rate of citrate reabsorption and metabolism in the proximal tubule. Reabsorption is mediated by an apical membrane 3Na+/citrate cotransporter encoded by the NaDC-1 gene, and metabolism is mediate by one of two pathways: a mitochondrial pathway that mediates citrate metabolism in the tricarboxylic acid cycle; and a cytoplasmic pathway in which ATP citrate lyase metabolizes citrate to acetyl CoA and oxaloacetate. The proposed studies will examine the molecular mechanisms responsible for regulation of citrate reabsorption and metabolism, focusing on NaDC-1. Studies in Aim 1 will address the molecular mechanisms of NaDC-1 regulation in rats with chronic metabolic acidosis, K+ deficiency, alkali feeding, and starvation. Studies in Aim 2 will further address the molecular mechanisms of NaDC-1 regulation in cultured proximal tubule cells exposed to acidic extracellular fluid. These studies will utilize cells expressing native NaDC-1, as well as cells expressing stably transfected tagged NaDC-1, and transiently transfected reporter constructs. Aim 1 and 2 together will address the regulation of NaDC-1. Studies in Aim 3 will address the role of signaling pathways known to be activated by acidosis in the regulation of NaDC-1. These pathways include: 1) tyrosine kinase and MAP kinase pathways; 2) glucocorticoids; and 3) endothelin. Studies in Aim 4 will quantitate citrate metabolism in mitochondrial and cytoplasmic pathways of the renal proximal tubule using 13C NMR spectroscopy. Lastly, studies in Aim 5 will address whether hyperkalemia causes hypercitraturia and thus is responsible for the lack of predisposition to nephrolithiasis in hyperkalemic renal distal tubular acidosis. These studies will allow us to continue to pursue an understanding of the mechanisms of regulation of renal citrate handling in the proximal tubule.